Differentiation potential of an immortalized non-tumorigenic human liver epithelial cell line as liver progenitor cells

Abstract

We report the differentiation potential of an immortalized non-tumorigenic human liver epithelial cell line, THLE-5b. Under basic culture conditions THLE-5b showed undifferentiated phenotypes. When grown as cell aggregates, THLE-5b exhibited a hepatocyte-like ultrastructure, ammonia metabolic activity and several other indicators that suggest hepatocytic maturation, including up-regulation or induction of liver-specific genes such as albumin and tryptophane 2,3-dioxygenase, and down-regulation of biliary cell markers such as gamma-glutamyl transpeptidase (GGT). Under these conditions, transcriptional factors such as HNF-1 and HNF-4alpha were also up-regulated or induced. In Matrigel culture, expression of GGT was up-regulated. THLE-5b expressed both albumin and alpha 1-antitrypsin, but lost expression of CK19 in severe combined immunodeficient mice. Thus, THLE-5b can be aligned with progenitor cells, which are committed to the hepatocytic or biliary epithelial cell lineage. These results imply that bipotent progenitor cell populations similar to THLE-5b cells may exist in adult human liver.

Presence of side-population cells in an immortalized nontumorigenic human liver epithelial cell line

“…THLE-5b is a nontumorigenic cell line (Lechner et al 1990), and THLE-5b SP cells are also nontumorigenic (no tumor formation occurred 6 mo after the inoculation of 5×10 6 SP cells/mouse into three nude mice). Previous results using the THLE-5b line implied that THLE-5b appears to represent liver progenitor cells and that a bipotent progenitor cell population similar to THLE-5b may exist in adult human liver (Tokiwa et al 2006). The current results support these possibilities.…”

“…However, whether stem/progenitor cells exist in the human liver has been controversial (Tokiwa et al 2006). Side-population (SP) cells have been shown to be highly enriched stem cells and are identified by flow cytometry on the basis of their unique ability to efflux the Hoechst 33342 dye (Goodell et al 1996).…”

“…Hepatic oval cells exhibit the SP phenotype (Shimano et al 2003), andHussain et al (2005) reported that SP cells derived from human livers possess the potential of hepatic stem cells. Our previous studies (Tokiwa et al 2006) showed that a SV40 T antigenimmortalized, nontumorigenic human liver cell line, THLE5b, appears to represent liver progenitor cells committed to the hepatocytic or biliary epithelial cell lineage. In this study, we used flow cytometry and staining with the Hoechst 33342 dye to investigate whether or not the THLE-5b line contains SP cells.…”

“…THLE-5b cells (Tokiwa et al 1998(Tokiwa et al , 2006 were supplied by Dr. CC Harris (NCI, Bethesda, MD), and HuH-7 and HepG2 were obtained from the Japanese Cancer Research Resource Bank. These cells were routinely cultured in Roswell Park Memorial Institute, Buffalo, New York (RPMI 1640) medium supplemented with 10% fetal calf serum (FCS) at 37°C in a humidified atmosphere containing 5% CO 2 and 95% air.…”

“…Reverse transcription-polymerase chain reaction (RT-PCR) was performed on mRNA isolated from both the SP and non-SP cells. Total RNA was isolated as described previously (Tokiwa et al 2006). The cDNAs were synthesized by first strand cDNA kit (ReverTraAce-α, Toyobo, Osaka, Japan).…”